RE: RTPCR from paraffin sections- still looking for that somebody!!

From:Luis Chiriboga

also recommend: American Journal of Pathology 158(2) FEB2001 419-429 K.
Specht et al "Quantitative gene expression analysis in microdissected
archival formalin-fixed and paraffin-embedded tumor tissue"

-----Original Message-----
From: Johnson, Teri []
Sent: Tuesday, May 06, 2003 9:45 AM
Subject: Re: RTPCR from paraffin sections- still looking for that


You might try this reference: J Histochem cytochem 50:1237-1245, 2002,
Uneyama et al.  They used methacarn fixed, paraffin embedded tissues for
their studies.

Teri Johnson
Managing Director Histology Facility
Stowers Institute for Medical Research
1000 E. 50th St.
Kansas City, Missouri  64110

-----Original Message-----
Sent: Tuesday, May 06, 2003 3:23 AM
Subject: R#233#f. : Re: RTPCR from paraffin sections- still looking for that

I'm a new subscriber and that was my first question, nice to have some
quick responses. Thanks Nina and Bob but I don't believe mRNA degradation
can explain our whole problem. Firstly (I didn't include this info in my
initial messsage) In situ hybridization with riboprobes works even better
in our paraffin sections (5u) than our frozen sections (15u), so in
paraffin sections we have 3 times less material but twice the signal. Its
true that PCR requires intact mRNA while ISH doesn't. Secondly we are also
unable to amplify genomic DNA from our paraffin sections. So we think its
more a question of digestion/extraction/accessibility of mRNA molecules.
We're now looking and testing extraction protocoles. We believe RTPCR from
paraffin sections is possible because the only three makers of
microdissection systems : LEICA, ARCTURUS, and PALM claim that it's
feasable. And thats why we bought the dam- !$#O*!?X ! $#  machine.
Somebody, Somebody out there must know how to do it.....

 truely yours,
Nancy Walker
Molecular Biology Scientist

Sanofi-Synthelbo Research
B.P. 37 Lab#233#ge Innopole
tel : (33)561004179#160# fax :(33)561004001

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